1. Field of the Invention
The present invention relates to a method and equipment for cultivating anaerobic ammonium-oxidizing bacteria. Particularly, the present invention relates to a method and equipment for cultivating anaerobic ammonium-oxidizing bacteria, which are intended for efficiently cultivating anaerobic ammonium-oxidizing bacteria that are employed for removing nitrogen from nitrogen containing liquids such as wastewater and anaerobically oxidize ammonium and nitrite, nitrogen components in the nitrogen containing liquids, used as substrates.
2. Description of the Related Art
Nitrogen components contained in nitrogen containing liquids such as sewage and industrial wastewater cause the eutrophication of water systems such as lakes and mashes and cause a reduction in dissolved oxygen in water systems. For these reasons, the nitrogen components must be removed before the nitrogen containing liquids are drained out of the systems. Principal nitrogen components contained in this nitrogen containing liquid are ammonium nitrogen, nitrite nitrogen, nitrate nitrogen, and organic nitrogen.
Conventionally, for treating this kind of wastewater, removal by an ion exchange method and oxidation with chlorine and ozone have been used for wastewater that contains nitrogen at low concentrations, while a method by biological treatment has been adopted for wastewater that contains nitrogen at moderate and high concentrations.
In the biological treatment, nitrification/denitrification treatment by aerobic nitrification and anaerobic denitrification is carried out. In the aerobic nitrification, ammonium nitrogen and nitrite nitrogen in a nitrogen containing liquid to be treated are aerobically oxidized with ammonium-oxidizing bacteria (such as the genera Nitrosomonas, Nitrosococcus, Nitrosospira, and Nitrosolobus) and nitrite oxidizing bacteria (such as the genera Nitrobactor, Nitrospira, Nitrococcus, and Nitospina) in a nitrification tank. On the other hand, in the anaerobic denitrification, anaerobic denitrification with heterotrophic bacteria such as Pseudomonas denitrificans is performed in a denitrification tank.
In the aerobic nitrification, the nitrification tank is operated at a load ranging from 0.2 to 0.3 kg-N/m3/d. On the other hand, in the anaerobic denitrification, the denitrification tank is operated at a load ranging from 0.2 to 0.4 kg-N/m3/d. Accordingly, when the total nitrogen concentration in the nitrogen containing liquid to be treated falls within the range of 30 to 40 mg/L, a retention time required is 6 to 8 hours for the nitrification tank and 5 to 8 hours for the denitrification tank, leading to a problem that each large-scale treatment tank must be provided.
In addition, the denitrification of nitrogen containing liquids that contain inorganic matter, such as industrial wastewater requires organic matter, although the loads of the nitrification and denitrification tanks are set to the above range. Thus, methanol must be added at a concentration 3 to 4 times greater than a nitrogen concentration in the nitrogen containing liquid to be treated, leading to another problem that large running cost in addition to initial cost is required.
By contrast, nitrogen removal by an anaerobic ammonium oxidation method is recently receiving attention, as disclosed in Japanese Patent Application Laid-Open No. 2001-037467. This anaerobic ammonium oxidation method is a method in which ammonium and nitrite in a nitrogen containing liquid to be treated are used as an electron donor and an electron acceptor, respectively, and are simultaneously denitrified with a group of anaerobic ammonium-oxidizing bacteria according to Chemical Formula 1 (see Strous M. et al., (1998) The sequencing batch reactor as a powerful tool for the study of slowly growing anaerobic ammonium-oxidizing microorganisms. Appl. Microbiol. Biotechnol, 50, 589-596) shown below.NH4++1.32NO2+0.066HCO3+0.13H+→1.02N2+0.26NO3+0.066CH2O0.5N0.15+2.03H2O  [Chemical Formula 1]
According to the anaerobic ammonium oxidation method, the use of methanol and so on required in conventional denitrification methods can be reduced significantly, because ammonium in the nitrogen containing liquid to be treated is utilized as a hydrogen donor. Additionally, this method also has a merit of reducing the amount of sludge generated by the treatment and as such, is considered to be effective as a future nitrogen removal method.
Microorganisms of the genus Planctomycetes have been reported as those responsible for this anaerobic ammonium oxidation reaction (Strous M. et al., (1999) Missing lithotroph identified as new planctomycete. Nature, 400, 446-449).
In general, the growth rates of microorganisms utilized in the above-described nitrogen removal treatment of wastewater are slow, and the yields of their bacterial cells are also low. Thus, the wastewater treatment requires a large amount of bacterial cells of those microorganisms and therefore presents a problem that the cultivation and acclimatization of the microorganisms require a great deal of time.
For dealing with this problem, Japanese Patent Application Laid-Open No. 09-187272 has proposed a method in which a nitrate bacterium with a slow growth rate is efficiently cultivated by controlling the amount of a substrate ammonium added in the logarithmic growth phase of the nitrate bacterium.
The anaerobic ammonium oxidation method is difficult to put to practical use and is not prevalent, in spite of the fact that a large number of anaerobic ammonium oxidation methods have been proposed.
One reason is that the growth rates of anaerobic ammonium-oxidizing bacteria used in the anaerobic ammonium oxidation method are slow, and the yields of the bacterial cells are low. Especially, microorganisms of the genus Planctomycetes have been reported to require a doubling time (a time when 1 bacterial cell is grown into 2 bacterial cells) of 11 days. This slow growth is a major challenge to the practical use of the method. Under present circumstances, a method for cultivating these anaerobic ammonium-oxidizing bacteria is not yet reported concretely.
Seed sludge that contains anaerobic ammonium-oxidizing bacteria is required for operating facilities that treat wastewater by the anaerobic ammonium oxidation method. Seed sludge with a large number of bacterial cells must be introduced for starting up a wastewater treatment equipment with stability within a shot time.
However, the efficient cultivation of bacteria requiring a doubling time as long as 11 days as seed sludge is a major challenge to the practical use of wastewater treatment by the anaerobic ammonium oxidation method. The practical use of the anaerobic ammonium oxidation method is in urgent need of overcoming this challenge.
Furthermore, the anaerobic ammonium oxidation method is capable of high-speed treatment as described above. Therefore, the method requires a consumption rate of a substrate as never experienced in cultivating nitrifying bacteria as disclosed in Japanese Patent Application Laid-Open No. 09-187272. In addition, the method requires a long cultivation period because of the slow growth rates of the bacteria and therefore requires an enormous amount of the substrate at the time of cultivation. Thus, the method presents additional problems associated with large cost required for feeding the substrate as well as large cost required for treating a large amount of waste liquids generated by cultivation.
The present invention has been achieved in light of such circumstances, and an object of the present invention is to provide a method and equipment for cultivating anaerobic ammonium-oxidizing bacteria, which can feed substrates without waste to generate seed sludge with high bacterial cell concentrations and can start up operation within a short time, in cultivating anaerobic ammonium-oxidizing bacteria with ammonium and nitrite as substrates.